Antonella Geromin

Autologous stem cell transplantation for systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is responsive to treatment with immunosuppressives and steroids, but often pursues a relapsing or refractory course resulting in increasing incapacity and reduced survival. Autologous stem cell transplantation (ASCT) following immunoablative chemotherapy is a newer therapy for autoimmune disease of potential use in severe SLE. A retrospective registry survey was carried out by the European Blood and Marrow Transplant and European League Against Rheumatism (EBMT/EULAR) registry. Data was collected from 53 patients with SLE treated by ASCT in 23 centres. Disease duration before ASCT was 59 (2-155) months (median, range), 44 (83%) were female, and median age was 29 (9-52) years. At the time of ASCT a median of seven American College of Rheumatology (ACR) diagnostic criteria for SLE were present (range 2-10) and 33 (62%) had nephritis. Peripheral blood stem cells were mobilized with cyclophosphamide and granulocyte colony stimulating factor in 93% of cases. Ex vivo CD34 stem cell selection was performed in 42% of patients. Conditioning regimens employed cyclophosphamide in 84%, anti-thymocyte globulin in 76% and lymphoid irradiation in 22%. The mean duration of follow-up after ASCT was 26 (0-78) months. Remission of disease activity (SLEDAI < 3) was seen in 33/50 (66%; 95%CI 52-80) evaluable patients by six months, of which 10/31 (32%; 95%CI 15-50) subsequentlyrelapsed after six (3-40) months. Relapse was associated with negative anti-double stranded DNA (anti-dsDNA) antibodies before ASCT (P 0.007). There were 12 deaths after 1.5 (0-48) months, of which seven (12%; 95%CI 3-21) were related to the procedure. Mortality was associated with a longer disease course before ASCT (P 0.036). In conclusion, this registry study demonstrates the efficacy of ASCT for remission induction of refractory SLE, although mortality appeared high. The safety of this procedure is likely to be improved by patient selection and choice of conditioning regimen. The return of disease activity in one-third of patients might be reduced by long-term immunosuppressive therapy post-ASCT.

Incidence, outcome, and risk factors of late-onset noninfectious pulmonary complications after unrelated donor stem cell transplantation

Summary:We evaluated the incidence, the risk factors, and the outcome of late-onset noninfectious pulmonary complications (LONIPCs) among 50 patients who underwent allogeneic stem cell transplantation from unrelated donors. Of the 39 patients surviving at least 3 months, 10 (26%) fulfilled the diagnostic criteria of LONIPCs and were further subclassified as having bronchiolitis obliterans (four patients), bronchiolitis obliterans with organizing pneumonia (four patients), and interstitial pneumonia (two patients). Two patients had a durable partial remission after treatment with prednisone and cyclosporine; the remaining eight patients did not respond to treatment and five of them died of respiratory failure. Advanced stage of disease at transplant and chronic extensive graft-versus-host disease (GVHD) were significantly associated with the development of LONIPCs. Pulmonary function test (PFT) results before transplantation were similar in all patients, but patients with LONIPCs had a significant decrease in PFT indexes at the third month after BMT compared with controls. Moreover, the rate of cyclosporine taper during the fourth and fifth months after BMT was significantly more rapid in patients with LONIPCs than in controls, suggesting that the risk of LONIPCs may be influenced by a faster reduction of GVHD prophylaxis.

Overexpression of multidrug resistance‐associated p170‐glycoprotein in acute non‐lymphocytic leukemia

Abstract: Resistance to several cytotoxic agents, including anthracyclines, vinca alkaloids and epipodophylline derivatives (multidrug resistance, or MDR) can develop in tumor cells by overexpression of a 170‐kd glycoprotein (p170) which is an essential component of a membrane transport system leading to increased drug efflux and decreased intracellular drug concentration. By means of a p170‐directed monoclonal antibody (MRK‐16) and immunocytochemistry (alkaline phosphatase anti‐alkaline phosphatase technique), we investigated the expression of p170 in marrow blast cells of 59 cases (38 at diagnosis and 21 in relapse) of acute‐non‐lymphocytic leukemia (ANLL). The proportion of strongly MDR‐positive cells was higher in relapse that at diagnosis (median 15.5% vs 1.5%). Out of 31 patients who were evaluable for the results of first remission induction, failure of first‐line treatment (including Daunorubicin, standard‐dose and high‐dose Arabinosyl Cytosine, and sometimes also Mitoxantrone) occurred in 8/22 MDR‐positive cases and in 1/9 MDR‐negative ones (p = 0.21). Failure of first‐line treatment was always associated with a progressive increase of p170 expression. Total failures (no remission plus early relapse) were more frequent (p = 0.001) among MDR‐positive cases (16/22) than among the others (2/9). These data show that MDR is very frequent in ANLL also at diagnosis and suggest that MDR can contribute to early failure of standard treatment.

P-glycoprotein (PGP), lung resistance-related protein (LRP) and multidrug resistance-associated protein (MRP) expression in acute promyelocytic leukaemia

We analysed the expression of three drug transporter proteins [p‐glycoprotein (PGP), lung resistance‐related protein (LRP) and multidrug resistance‐associated protein (MRP1)] involved in anthracycline resistance that are frequently overexpressed in poor‐risk adult acute non‐lymphocytic leukaemia (ANLL), in 23 acute promyelocytic leukaemia (APL) patients at onset managed at a single institution. Cellular daunorubicin accumulation was also evaluated. At onset, no case had PGP or MRP1 expression that exceeded that of non‐multidrug‐resistant (MDR) cell lines. Only one case showed LRP overexpression. No peculiar MDR features distinguished the seven patients who relapsed from those who maintained complete remission. In the onset vs. first relapse, only one patient showed an increased (threefold) PGP expression at relapse. At second relapse, three out of four patients showed a PGP expression two‐ to threefold higher than baseline values. These results are consistent with the view that low PGP, LRP and MRP1 expression and the absence of defects in intracellular drug accumulation may account for the peculiarly high sensitivity of APLs to anthracycline. It does not support the screening of MDR markers in APL patients at onset as predicting factors of early relapse. The results suggest that no significant changes in PGP, LRP or MRP1 expression are likely to occur at first relapse. In contrast, PGP expression is likely to increase later in the patient history as a result of additional chemotherapy courses.

P-glycoprotein, lung resistance-related protein and multidrug resistance-associated protein in de novo adult acute lymphoblastic leukaemia

Summary.  P‐glycoprotein (P‐gp), lung resistance‐related protein (LRP) and multidrug resistance‐associated protein (MRP) expression, and blast cell intracellular daunorubicin accumulation (IDA) were evaluated in 95 previously untreated cases of adult acute lymphoblastic leukaemia (ALL) using flow cytometry. Forty‐five out of 95 (47%) patients were P‐gp positive (+), 12/66 (18%) were LRP+ and 11/66 (17%) were MRP+. Eighteen out of 66 (28%) patients showed a simultaneous multidrug resistance (MDR)‐related protein expression higher than controls for more than one protein, while 24/66 (36%) cases did not overexpress any protein. Twenty‐one out of 24 (87%) cases overexpressing at least one MDR‐related protein had a defect in accumulating daunorubicin into their blast cells, while only 4/24 (16%) cases who did not overexpress any protein had similar features. The complete remission rates were similar in MDR‐positive and ‐negative (–) patients but relapses within 6 months were more frequent in P‐gp+ cases, and therefore the disease‐free survival duration was shorter in P‐gp+ than in P‐gp– patients (P = 0·01). The number of MRP+ and/or LRP+ cases was too small to be able to draw any conclusion on their role in affecting or predicting therapy outcome. In conclusion, P‐gp overexpression associated with a defect in daunorubicin accumulation is a frequent feature in adult ALL at onset and seems to be related to poorer therapy outcome and, consequently, a shorter disease‐free survival. LRP and MRP overexpression seems to be a rare event and no conclusion can be drawn on its prognostic role.

Fludarabine-based induction therapy does not overcome the negative effect of ABCG2 (BCRP) over-expression in adult acute myeloid leukemia patients

Over-expression of multidrug resistance (MDR) proteins PGP and BCRP has a negative prognostic impact in acute myeloid leukemia (AML) patients. Inclusion of fludarabine in induction chemotherapy increases remission rate in PGP over-expressing cases. We investigated the role of BCRP in 138 adult AML patients receiving induction therapy with fludarabine. None of the MDR-related proteins influenced complete remission attainment. Conversely, high levels of BCRP significantly affected disease-free survival, as higher relapse rates (48.5% vs 28.5%) and earlier relapse occurred in BCRP+ patients. Also overall survival was affected by BCRP positivity, and survival significantly worsened in case of concomitant PGP and BCRP over-expression.

Dendritic cell recovery after autologous stem cell transplantation

There is persistent immunosuppression not only in allogeneic but also in autologous stem cell transplantation because humoral and cellular immunity may take a year or more to return to normal, with increased risk of infectious complications. This immune defect may also involve antigen presentation, in particular dendritic cell (DC) function. We evaluated DC subset reconstitution in 58 patients who underwent bone marrow (BM) or peripheral blood (PB) autologous haematopoietic stem cell transplantation (HSCT). In all patients DC type 1 (DC1) and DC type 2 (DC2) were already significantly lower than in normal individuals before conditioning therapy (DC1/μl 3.1 ± 1.0, DC2/μl 3.0 ± 1.1). On day 0 and day +7 the mean DC1 and DC2 numbers were very low in both groups. Patients who received unmanipulated marrow or peripheral blood stem cells reached pre-conditioning levels of DC1 and DC2 cells on day +20. In patients receiving selected CD34 cells, DC increased slowly and pre-transplant counts were observed only on day +60. Nearly ‘normal’ levels of DC1 and DC2 could be observed in the first group from day +180, and were maintained thereafter; in CD34+ selected patients DC1 and DC2 counts remained lower than normal. Our data emphasise that circulating antigen presenting cells (APC) recover quickly. It remains to be determined if DC frequency in PB reflects their tissue function. The relatively low incidence of infections in patients undergoing autologous transplantation, despite defective lymphocyte reconstitution, could be related to functionally efficient DC.

Clinical characteristics, prognostic factors and multidrug‐resistance related protein expression in 36 adult patients with acute promyelocytic leukemia

Abstract: We report on a single‐center experience about the characteristics and outcome of 36 acute promyelocytic leukemia (APL) patients observed at our Department of Hematology between 1990 and 2002. The expression, of multidrug‐resistance (MDR) associated proteins (PGP, LRP, MRP1) was also analyzed. There were 12 males and 24 females (median age 37 yr), 89% (32 of 36) with classic morphology, and 11% (four of 36) with a microgranular variant. Risk class (according to GIMEMA/PETHEMA): 25% (nine of 36) high risk (HR), 53% (nineteen of 36) intermediate risk (IR), 22% (eight of 36) low risk (LR). PGP, LRP, and MRP1 expression at onset and at first relapse was low. CD33 antigen expression was high in all cases. The patients were treated according to GIMEMA protocols (LAP0389 and AIDA) including ATRA in induction in 75% (27 of 36) of cases and 94% (34 of 36) achieved a complete remission (CR) after induction therapy while 6% (two of 36) died early (DDI) of hemorrhage. Outcome: 71% (24 of 34) of evaluable patients remain in CR at a median follow‐up of 57 months (range 4–158 months) while 29% (10 of 34) relapsed at a median time of 12 months (range 8–43 months) and, of them, eight of 10 died early. The majority of patients that relapsed were in high‐risk group. The overall survival (OS) of the whole population at 32 months was 66% and the DFS at 42 months was 62%. A statistically significant difference in terms of DFS was observed between HR and IR/LR patients (P = 0.04 by log‐rank). DFS was not affected by age, sex, Hb levels, karyotype, and BCR isoform. At conclusion, our data confirm that despite the high rate of success with ATRA plus chemotherapy as induction (more than 90% of CR), about 30% of APL patients have a relapse (without a long‐lasting second remission) and underline the importance of patient stratification in distinct risk groups at diagnosis in order to better adapt the type and intensity of treatment (risk‐adapted therapy). Taking into account the high expression of CD33 and the low expression of MDR proteins in APL, new and investigational approaches like gemtuzumab–ozogamicin, with or without ATRA and other new drugs, should be strongly considered expecially in HR APL.

Prognostic factors and outcome of Epstein–Barr virus DNAemia in high-risk recipients of allogeneic stem cell transplantation treated with preemptive rituximab

This study assessed incidence, predictive factors, and outcome of Epstein–Barr virus (EBV) DNAemia in 100 recipients of allogeneic hematopoietic stem cell transplant. A total of 68 patients received anti‐thymocyte globulin before unrelated grafts.

mdr-1 GENE AMPLIFICATION IN ACUTE LYMPHOBLASTIC LEUKAEMIA PRIOR TO ANTILEUKAEMIC TREATMENT

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